JCS02708 Supplementary Material

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• Supplemental Figure 1 -

  Fig. S1. Density-dependent generation of p75^NGFR+ smooth muscle and glia. E14.5 rat cortical cells at passage 1 were plated at medium density, expanded in FGF2 ± BMP2 for 3 days, then differentiated ± BMP2 for an additional 5 days. The resulting colonies contained a dense core and a sparse periphery, as previously described (Rajan et al., 2003). (A-B) Low magnification image of colony shows intense GFAP (green) staining in the dense core of both FGF2-only (A) and FGF2/BMP2 (B) cultures, although the GFAP⁺ cells are flatter in FGF2/BMP2 co-treated cultures. SMA⁺ (red) cells were absent in the FGF2-only culture (A) but abundant in the sparse edge of the FGF2/BMP2 co-treated culture. The two cell types occupied mutually exclusive locations based on local density. (C,D) While no SMA⁺p75^NGFR+ cells were generated from FGF2-expanded/withdrawn cells (C), FGF2/BMP2-treated cultures generated some SMA⁺ cells that retained p75^NGFR expression (D), consistent with its co-expression in a subset of CPm cells in vivo. (E,F) While FGF2-expanded/withdrawn cultures generated only immature GFAP⁺p75^NGFR− CNS astrocytes (E), a large proportion of the BMP2-generated flattened GFAP⁺ cells co-expressed p75^NGFR (F). (G,H) Quantitation shows mean ± s.e.m., n=3-4. Bar for A,B=80 mm; C-F=20 mm.